Specific Challenge Topic 14-DK-101, "Induced Pluripotent stem cells-cellular and humanized mouse" Somatic cells such as fibroblasts from patients with diseases can be used to create cell lines, tissues, and perhaps, organ systems, through induced pluripotent stem cell (iPSC) technology. Such models could be used to elucidate underlying pathology of disease or screen for agents that could be used therapeutically. Combining this approach with mouse strains able to accept multiple human tissues without rejection could provide the microenvironment milieu to support the tissue's physiological function within the context of the whole organism, enabling greater understanding of disease pathogenesis and providing a platform for preclinical testing of drug candidates." The liver performs over 3,000 functions critical to maintaining the organism, ranging from production and secretion of proteins such as albumin, clotting factors and antiproteases, to metabolism and excretion of exogenous compounds such as drugs or toxins as well as endogenous compounds like hormones, bilirubin and bile acids and cholesterol. Liver-based metabolic disease can result from mutations in genes in these critical pathways. Transgenic or knock-out mouse models have been created for many of these diseases, but they do not always faithfully reproduce the human disease. For example mutations in the bile salt export pump (BSEP) results in severe cholestasis and fibrosis in human patients requiring whole organ transplants, yet the knockout mouse is nearly without a phenotype. While many patients with alpha-1-antitrypsin (A1AT) deficiency develop liver fibrosis/cirrhosis many times resulting in liver transplantation, the transgenic mouse model carrying mutant human genes shows an extremely mild phenotype. The mouse model for a deficiency of ornithine transcarbamylase (OTC) activity, the rate limiting step in ammonia metabolism, is a fair model, however the animals tolerate a diet containing normal amounts of protein, while severely affected human patients require severe protein restriction to prevent lethal hyperammonemia. Mice (and rats) are particularly poor models for studying liver fibrosis and cirrhosis, common features of many human liver diseases. Humanized mice may offer a platform to both the study and treatment of hepatic fibrosis and cirrhosis. These are only a few examples of mouse models that do not faithfully recreate the human disease. We propose, the hypothesis, that the best models for human metabolic liver disease are those created from the affected human hepatocytes. Thus, we propose to" humanize" the liver of FRG mice by transplantation of affected human hepatocytes to create authentic models of human metabolic disease. These mice are immunodeficient and also deficient in the tyrosine catabolic enzyme, fumarylacetoacetate hydrolase (Fah -/-) and develop irreversible liver failure if left untreated. However, if Fah-proficient cells are transplanted, they readily and rapidly repopulate the native liver with donor cells, even if the donor cells are of human origin (a). To create these models, the liver of FRG mice will be "humanized" with hepatocytes derived from patients with metabolic disease. In addition, iPSC technology will be utilized to reprogram the liver cells from metabolic disease patients and following hepatic differentiation, additional mice will be humanized with iPS-derived hepatocytes. These humanized mouse models can then be compared to the authentic diseased liver with respect to changes in clinical chemistry (of the patient or animal), the histopathology of the liver and gene and protein expression profiling of liver tissue. In this manner, we will be able to determine if the humanized models developed with these procedures faithfully reproduce the phenotype observed in the patient. In addition to the direct effects on metabolic liver disease, success with these models will facilitate the use of humanized mouse models to investigate other liver based diseases such as Wilson's and Alpha-1-antitrypsin deficiency and may even lead to better-humanized models for primary, acute liver failure and viral, alcoholic and autoimmune hepatitis and even the hepatic stage of malaria, especially if a human immune system were reconstituted in addition to the liver. PUBLIC HEALTH RELEVANCE: Genetic defects in liver functions affect approximately 1/50,000 live births. For most, there are no small animal models that faithfully reproduce the human phenotype. We propose the hypothesis that the most appropriate model for these liver based diseases is one that is produced with authentic disease-affected hepatocytes. We have recently succeeded in humanizing the liver of specialized mice by repopulating the liver with human hepatocytes. We propose to develop models of human liver disease by transplantation of liver cells from patients with metabolic liver diseases and characterize the model for relevance to the human disease. In addition we will reprogram the liver cells to produce induced pluripotent cells (iPSC) that can also be used to repopulate and humanize the mouse liver. We propose these humanized mice will be a technology platform to better understand the human disease and it will aid in the development of gene or cellular therapy to correct these devastating diseases.